Insecticidal crystal proteins (encoded by cry genes) produced by Bacillus thuringiensis (Bt) are fatal for insects of different orders such as Lepidoptera. The genes that encoded these crystal proteins can be detected on plasmids and chromosomal DNA and show different types in various strains. Therefore, the objective of this study was to determine molecular characteristics of Iranian Bt strains as well as their toxicity against Ephestia kuehniella (Zeller) (Lepidoptera: Pyralidae). The collection sites included fields, gardens, and desert and semi-desert areas in 8 provinces of Iran. For crystal formation, each isolate was cultured in T3 medium. The results showed that the Bt isolates produced different types of crystals including spherical (73.33%), bipyramidal (53.33%), irregular (40%), cubical (33.33%), and elliptical (13.33%). Plasmid DNA extraction was performed and showed that most of the strains exhibited similar pattern in the number and the size of the plasmid bands to those of Bt kurstaki (Btk). Some specific primers were used for PCR amplification to distinguish different crystal genes including cry1 (A, C, and D) and cry2A (a and b). The primers related to cry1D and cry2Aa1 genes produced no amplicons. The results revealed that the most abundant gene was cry1-type. All strains analyzed for the cry2Ab2 gene presented unexpected bands. Electrophoretic profile of the protein crystals showed bands with different diversity in number, and size ranged from about 16 to 140 KDa. The bioassay result of some more toxic strains exhibited that the pathogenicity of 1019 was higher than the rest, even the reference strain, Btk. However, the toxicity of other strains was the same as Btk. According to the phylogenetic tree, 1019 was located in the same group with Bt subspecies coreanensis, Bt subspecies indiana and Bt subspecies tolworthi. The investigated Iranian strains had the lepidopteran-active cry genes. The strains with the same toxicity to E. kuehniella had various cry genes, plasmid, and crystal protein profiles and vice versa. Therefore, characterization of cry genes of native strains could lead to access potent isolates as biocontrol agents against native insect pests.

中文翻译：

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伊朗苏云金芽孢杆菌菌株的分离和鳞翅目活性cry基因的表征

苏云金芽孢杆菌（Bt）产生的杀虫晶体蛋白（由cry基因编码）对鳞翅目等不同目昆虫是致命的。编码这些晶体蛋白的基因可以在质粒和染色体 DNA 上检测到，并在各种菌株中显示出不同的类型。因此，本研究的目的是确定伊朗 Bt 菌株的分子特征以及它们对 Ephestia kuehniella (Zeller)（鳞翅目：Pyralidae）的毒性。收集地点包括伊朗8个省的田野、花园、沙漠和半沙漠地区。对于晶体形成，每个分离物都在 T3 培养基中培养。结果表明，Bt分离株产生了不同类型的晶体，包括球形（73.33%）、双锥（53.33%）、不规则（40%）、立方（33.33%）和椭圆形（13.33%）。进行质粒 DNA 提取并显示大多数菌株在质粒条带的数量和大小上表现出与 Bt kurstaki (Btk) 相似的模式。一些特异性引物用于PCR扩增以区分不同的晶体基因，包括cry1（A、C和D）和cry2A（a和b）。与cry1D 和cry2Aa1 基因相关的引物没有产生扩增子。结果表明，最丰富的基因是cry1型。对cry2Ab2 基因进行分析的所有菌株都呈现出意料之外的条带。蛋白质晶体的电泳图谱显示出数量不同的条带，大小范围从约 16 到 140 KDa。一些毒性较大的菌株的生物测定结果表明，1019的致病性高于其他菌株，即使是参考菌株Btk。然而，其他菌株的毒性与Btk相同。根据系统发育树，1019与Bt亚种coreanensis、Bt亚种indiana和Bt亚种tolworthi位于同一组。被调查的伊朗菌株具有鳞翅目活性的哭声基因。对E. kuehniella 具有相同毒性的菌株具有不同的cry 基因、质粒和晶体蛋白谱，反之亦然。因此，对天然菌株的cry 基因的表征可以导致获得有效的分离株作为针对天然害虫的生物防治剂。kuehniella 具有各种cry 基因、质粒和晶体蛋白谱，反之亦然。因此，对天然菌株的cry 基因的表征可以导致获得有效的分离株作为针对天然害虫的生物防治剂。kuehniella 具有各种cry 基因、质粒和晶体蛋白谱，反之亦然。因此，对天然菌株的cry 基因的表征可以导致获得有效的分离株作为针对天然害虫的生物防治剂。